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An oxidant and organic solvent tolerant alkaline lipase by P. aeruginosa mutant: downstream processing and biochemical characterization.

Authors
  • Bisht, Deepali1
  • Yadav, Santosh Kumar1
  • Darmwal, Nandan Singh1
  • 1 Centre of Excellence, Department of Microbiology, Dr. Ram Manohar Lohia Avadh University, Faizabad, U.P., India. , (India)
Type
Published Article
Journal
Brazilian journal of microbiology : [publication of the Brazilian Society for Microbiology]
Publication Date
Dec 01, 2013
Volume
44
Issue
4
Pages
1305–1314
Identifiers
PMID: 24688527
Source
Medline
Keywords
License
Unknown

Abstract

An extracellular alkaline lipase from Pseudomonas aeruginosa mutant has been purified to homogeneity using acetone precipitation followed by anion exchange and gel filtration chromatography and resulted in 27-fold purification with 19.6% final recovery. SDS-PAGE study suggested that the purified lipase has an apparent molecular mass of 67 kDa. The optimum temperature and pH for the purified lipase were 45 °C and 8.0, respectively. The enzyme showed considerable stability in pH range of 7.0-11.0 and temperature range 35-55 °C. The metal ions Ca(2+), Mg(2+) and Na(+) tend to increase the enzyme activity, whereas, Fe(2+) and Mn(2+) ions resulted in discreet decrease in the activity. Divalent cations Ca(+2) and Mg(+2) seemed to protect the enzyme against thermal denaturation at high temperatures and in presence of Ca(+2) (5 mM) the optimum temperature shifted from 45 °C to 55 °C. The purified lipase displayed significant stability in the presence of several hydrophilic and hydrophobic organic solvents (25%, v/v) up to 168 h. The pure enzyme preparation exhibited significant stability and compatibility with oxidizing agents and commercial detergents as it retained 40-70% of its original activities. The values of K(m) and Vmax for p-nitrophenyl palmitate (p-NPP) under optimal conditions were determined to be 2.0 mg.mL(-1) and 5000 μg.mL(-1).min(-1), respectively.

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